Cryptococcus neoformans is an opportunistic yeast that may produce a life-threatening meningitis in individuals with deficiencies in cell mediated immunity, including AIDS. The yeast is surrounded by a polysaccharide capsule that is essential to virulence. The primary constituent of the capsule is a polysaccharide termed glucuronoxylomannan (GXM). The capsule's ability to inhibit phagocytosis is its most important function. The overall goal of this study is to understand the cellular and molecular basis for inhibition of phagocytosis by the capsule. A library of GXM mAbs was generated during the current grant period. Depending on epitope specificity, a mAb can be highly opsonic or poorly opsonic. Such opsonization is not Fc dependent. Additional studies have found that the capsule is a matrix with a density that decreases in a near linear fashion as the capsule edge is approached. It is the overall hypothesis for this proposal that the position of binding of potentially opsonic ligands, e.g., antibody or opsonic fragments of C3 relative to the capular edge and the mobility of such ligands will determine whether or not the yeast will be ingested by phagocytes. Specific Aim 1 will identify, at a molecular level, the antibody-capsule interactions that distinguish highly opsonic from weakly opsonic mAbs. Aim 2 will assess the spatial relationship between GXM at the capsular edge and potentially opsonic ligands that are bound to the capsular matrix. Aim 3 will examine the molecular mobility of GXM and potentially opsonic ligands at the capsular edge and the effects of highly opsonic and weakly opsonic mAbs on such mobility. Aim 4 will identify structural components of GXM that are required for direct binding to macrophages. Aim 5 will identify means to activate macrophages for increased uptake of unopsonized and iC3b-opsonized cryptococci. These studies will contribute to our understanding of a virulence factor that is common to many microbial pathogens and will identify means to enhance phagocytosis and host resistance to cryptococcosis.